Ionotropic glutamate receptors activate ligand-gated cation channels that mediate the predominant component of excitatory neurotransmission in the central nervous system (CNS). These receptors have been classified based on their preference for the glutamate-like agonists (RS)-2-amino-3-(3-hydroxy-5-methyl-4-isoxazolyl)propionic acid (AMPA), kainate (KA), and N-methyl-D-aspartate (NMDA). All three glutamate receptor subtypes are heteromultimeric complexes, and many of the subunits that comprise them have been identified and characterized. To date, six NMDA receptor subunits (NR1, NR2A-2D and NR3A) have been reported.
The NMDA receptor (NMDAR) has unique properties distinguishing it from the other glutamate receptor subtypes. First, the activation of NMDAR requires the presence of dual agonists, glutamate (or NMDA) and glycine. The ligand-gated ion channel of the NMDA receptor is, thus, under the control of at least two distinct allosteric sites. In addition, the NMDA receptor controls the flow of both divalent (Ca.sup.2+) and monovalent (Na.sup.+, K.sup.+) ions into the postsynaptic neural cell through a receptor associated channel. (Foster et al., “Taking apart NMDA receptors”, Nature, 329:395-396, 1987; Mayer et al., “Excitatory amino acid receptors, second messengers and regulation of intracellular Ca.sup.2+ in mammalian neurons,” Trends in Pharmacol. Sci., 11:254-260, 1990). The activation of these receptors is regulated by Mg.sup.2+ in a voltage-dependent manner (i.e., the NMDAR is blocked at resting membrane potential and activated when depolarized). Most importantly; however, the NMDAR is extremely permeable to Ca.sup.2+, a key regulator of cell function.
NMDARs are believed to play a pivotal role in the transmission of excitatory signals from primary sensory neurons to the brain through the spinal cord (A. H. Dickenson (1990) Trends Pharmacol. Sci., 11. 307-309). NMDA receptors mediate Ca.sup.2+ influx into neurons, and its receptor-gated channel activity is blocked by Mg.sup.2+ in a voltage-dependent manner. These unique properties allow NMDARs to play a critical role in development of the nervous system, synaptic plasticity, memory, and other physiological processes in the CNS.
However, excessive stimulation of NMDARs has also been implicated in many pathological conditions including stroke, ischemia, head and spinal trauma, headache, epilepsy, neuropathic pain syndromes including diabetic neuropathy, glaucoma, depression and anxiety, drug addiction/withdrawal/tolerance, and in chronic neurodegenerative states, such as Alzheimer's disease, Huntington's disease, HIV-associated dementia, Parkinson's disease, multiple sclerosis, and amyotrophic lateral sclerosis (ALS).
The molecular cloning and functional analysis of expressed NMDAR subunits, coupled with the examination of their temporal and spatial expression patterns in vivo, has led to significant advances in our understanding of NMDAR function at the molecular level. However, the identification of these subunits alone has failed to explain the observed diversity in NMDAR function, particularly in motor neurons. Thus there is a need to further understand the role of NMDAR subunits in regulating these diverse functions.
Due to its broad-spectrum of neurological involvement, yet non-universal distribution, investigators are interested in the identification and development of drugs acting at the NMDA receptor. Drugs acting on the NMDA receptor are, therefore, expected to have enormous therapeutic potential. For instance, U.S. Pat. No. 4,904,681, issued to Cordi et al. (Cordi I), describes the use of D-Cycloserine, which was known to modulate the NMDA receptor, to improve/enhance memory and to treat cognitive deficits linked to a neurological disorder. D-Cycloserine is described as a glycine agonist which binds to the strychnine-insensitive glycine receptor.
U.S. Pat. No. 5,061,721, issued to Cordi et al. (Cordi II), describes the use of a combination of D-cycloserine and D-alanine to treat Alzheimer's disease, age-associated memory impairment, learning deficits, and psychotic disorders, as well as to improve memory or learning in healthy individuals.
U.S. Pat. No. 5,086,072, issued to Trullas et al., describes the use of 1-aminocyclopropanecarboxylic acid (ACPC), which was known to modulate the NMDA receptor as a partial agonist of the strychnine-insensitive glycine binding site, to treat mood disorders including major depression, bipolar disorder, dysthymia and seasonal effective disorder. It is also therein described that ACPC mimics the actions of clinically effective antidepressants in animal models. In addition, a copending U.S. patent application is cited that describes that ACPC and its derivatives may be used to treat neuropharmacological disorders resulting from excessive activation of the NMDA receptor.
None of the foregoing offers, however, a satisfactory mechanism for modulating NMDA receptor function. Development of drugs targeting the NMDA receptor, although desirous, has been hindered because the molecular pathway surrounding the NMDA receptor has not yet been completely elucidated. As mentioned above, the NMDAR consists of several protein chains (subunits) embedded in the postsynaptic membrane. Subunits NR1A and NR2A-D from a large extracellular region which probably contains most of the allosteric binding sites, several transmembrane regions looped and folded to form a pore or channel which is permeable to Ca.sup.2+, and a carboxyl terminal region. It is believed that the channel is in constant motion, alternating between a cation passing (open) and a cation blocking (closed) state. The opening and closing of the channel is regulated by the binding of various ligands to domains of the protein residing on the extracellular surface and separate from the channel. As such, these ligands are all known as allosteric ligands. The binding of two co-agonist ligands—glycine and glutamate—is thought to effect a conformational change in the overall structure of the protein which is ultimately reflected in the channel opening, partially opening, partially closing, or closing. The binding of other allosteric ligands modulates the conformational change caused or effected by glutamate and glycine. The recently characterized subunit NR3A has been found to act in a novel manner, as compared to subunits NR1A-2D. NR3A downmodulates the NMDAR and this downmodulation has been correlated with a decreased unitary current and Ca.sup.2+ permeability of the channel. This unique regulatory behavior associated with the NR3A subunit is believed to have therapeutic importance. For example, studies in mice have shown that the NR3A subunit may protect the young nervous system from excitotoxic damage during development. Thus, it is desirable to further understand the NR3A molecular pathway, thereby allowing for the discovery of therapeutic compounds that modulate this same pathway.